A kind of Bacillus xylosinolysine and its method for preparing α-keto acid

A technology of Bacillus glycolysine and Bacillus lysine, which is applied in the field of Bacillus xylosyllysine to prepare α-keto acid, which can solve the problem that the production intensity of α-keto acid is not high and it is difficult to realize industrialization problems such as broad substrate and product tolerance, broad substrate specificity, and less by-products

Active Publication Date: 2018-10-12
CHONGQING UNIV OF POSTS & TELECOMM
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although L-amino acid deaminase can specifically catalyze the deamination of L-amino acids without the generation of hydrogen peroxide, which avoids the further decarboxylation of α-ketoacids catalyzed by hydrogen peroxide, but the hi

Method used

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  • A kind of Bacillus xylosinolysine and its method for preparing α-keto acid
  • A kind of Bacillus xylosinolysine and its method for preparing α-keto acid

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Experimental program
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Effect test

Embodiment 1

[0027] Embodiment 1: strain screening

[0028] (1) 20 soil samples collected from different areas were naturally air-dried and crushed, and 5 g of crushed soil samples were added to conical flasks containing 50 ml of sterile water, shaken (200 rpm) at 30 ° C for 20 min, and statically Take 1ml of the supernatant for gradient dilution, and then spread 50μl of the diluted solution on a petri dish loaded with solid medium, and incubate at 30°C for 3 days. The solid medium used for enrichment is: peptone 10g / L, beef extract 3g / L, sodium chloride 5g / L, agar 15g / L, pH7.2.

[0029] (2) Single colonies grown on the solid medium in step (1) were picked respectively, inoculated into 5 ml sterile liquid medium, and cultured on a shaker (200 rpm) at 30° C. for 24 hours. Take 1ml of the bacterial liquid, and use the 2,4-dinitrophenylhydrazine method to detect pyruvate. The positive strain was inoculated onto the solid medium in step (1), cultured at 30°C for 24h, then inoculated into 5ml...

Embodiment 2

[0030] Embodiment 2: bacterial strain identification

[0031] The strain with the highest L-alanine oxidative deamination activity was identified by 16s rDNA. The strain has 100% homology with Lysinibacillus xylanilyticus, and it was named Lysinibacillus xylanilyticus XX-2. See the appendix for the 16s rDNA sequence.

Embodiment 3

[0032] Embodiment 3: Bacterial strain culture and thalline harvest

[0033] The slant medium is: 3g / L beef extract, 5g / L yeast extract, 5g / L sodium chloride, 15g / L agar, pH6.0.

[0034] The seed medium is: 3g / L beef extract, 5g / L yeast extract, 5g / L sodium chloride, pH6.0.

[0035] Enzyme production medium: 3g / L beef extract, 5g / L yeast extract, 5g / L sodium chloride, 3g / L L-alanine, pH6.0.

[0036] Slant culture: Inoculate the screened Bacillus xylosinus XX-2 on the slant medium, and culture at a constant temperature of 30°C for 48 hours;

[0037] Seed culture: inoculate the strains cultivated on the slant with an inoculation loop in 5ml of fermentation medium under aseptic conditions, and culture on a shaker (180rpm) at 30°C for 48 hours to obtain a seed solution;

[0038] Shake flask culture: the seed solution was introduced into fresh fermentation medium with 10% inoculum, and cultured on a shaker (180 rpm) at 30° C. for 48 hours.

[0039] Bacterial cell harvesting: the ...

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Abstract

The invention belongs to the technical field of biology and particularly relates to lysinibacillus xylanilyticus and a method for preparing alpha-ketoacid with the same. The lysinibacillus xylanilyticus XX-2 with a preservation number being CCTCC No:M2015520 is preserved in the China Center for Type Culture Collection. Whole cells of the lysinibacillus xylanilyticus XX-2 are taken as a biocatalyst to realize oxidative deamination of L-amino acids to obtain corresponding alpha-ketoacid, ammonia gas and hydrogen peroxide in the presence of air by L-amino acid oxidase in the lysinibacillus xylanilyticus XX-2, and in-situ decomposition of the generated hydrogen peroxide is realized by co-expressed catalase in the cells. The method overcomes defects of chemical synthesis methods and fermentation methods for preparing the alpha-ketoacid, has advantages of mild reaction conditions, environment friendliness, low cost, wide product spectrum and the like and is applicable to industrial production of the alpha-ketoacid.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a xylosyllysine bacillus and a method for preparing α-ketoacid by transforming L-amino acid with the strain. Background technique [0002] In organisms, α-keto acids are not only intermediates in sugar metabolism, but also participate in the biosynthesis of essential amino acids. α-keto acids are a class of bifunctional compounds, and are key intermediates in organic synthesis, drug synthesis and biosynthesis, and are widely used in biomedicine, food, cosmetics, feed and other fields. For example, four α-ketoacid calcium salts (α-ketoisocaproate calcium, α-ketoisovalerate calcium, α-keto-β-methylvalerate calcium, phenylpyruvate calcium), 1 hydroxyacid calcium salt (racemic hydroxymethionine calcium), a compound of 5 kinds of L-amino acids (L-lysine acetate, L-threonine, L-tryptophan, L-histidine, L-tyrosine) α-ketoacid tablets (Kaitong) are used to prevent and treat dam...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12P7/40C12P7/50C12R1/01
Inventor 夏仕文熊文娟韦燕婵何从林徐红梅
Owner CHONGQING UNIV OF POSTS & TELECOMM
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